Alzheimer's disease is predicted to be one of the highest priority global health risks in coming decades, yet there are currently no effective prophylactic or disease modifying therapies. The primary focus in the field has been to target reduction of the synaptotoxic amyloid ?-peptide (A?) however, recent late-stage clinical trials have proven disappointing. Therefore, new strategies and cellular targets are required. Recent work has found that genetic variants of genes involved in endocytosis such as Bridging Integrator 1 (BIN1 or amphiphysin 2/AMPH2) and phosphatidylinositol-binding clathrin assembly protein (PICALM) correlate with AD risk. Using bioinformatics analysis of STRING Consortium, gene neighborhood, gene fusions, gene co-occurrence, co- expression, experimental data, databases, text mining and protein homology indicate BIN1 is predicted to interact functionally with PICALM and critically, Synaptojanin1 (Synj1). Synj1 is a lipid phosphatase enriched in brain which dephosphorylates the critical signaling lipid phosphatidylinositol-4,5-bisphosphate [P(4,5)P2]. Phosphoinositides, known to be critical for neuronal function, have been shown by our group and others to be altered in AD affected patient brain as well as in mouse models. Synj1 is central to PI(4,5)P2 maintenance at the synapse and we have shown that loss of one copy of Synj1 has been shown to abrogate A?-induced synaptic dysfunction. AD and DS show similar pathologies in which APP processing and A? is central. DS individuals are trisomic for APP and Synj1, and Synj1 regulatory kinase DYRK1A and are at high-risk for developing AD. Synj1 has been shown to modify endosomal dysfunction in DS mouse models. Current studies have determined that iPSC derived neurons from DS individuals share similar pathogenic hallmarks of AD including increased A?42 production and decreased A?40:A?42 ratio. However, neuronal differentiation displayed normal neuronal conversion kinetics though one study observed exaggerated glia production, a phenotype associated with DS. Therefore, we hypothesize that Synj1 may be central to modifying synaptic and endosomal pathologies in AD and DS. We will test the hypothesis that aberrant Synj1 expression and function in neuronal phenotypes can recapitulate endosomal dysfunction in human induced pluripotent stem cell (iPSC) derived neurons. iPSC from sporadic AD and DS will be differentiated into forebrain neurons. Phenotypes shown to be mediated by Synj1 will be recapitulated in functional human neurons including endosome morphology and function, spine morphology, lipid dysregulation and functional complex formation with BIN1, PICALM, and Dyrk1a. CRISPR/Cas9 will be used to knock-out Synj1 alleles in AD and DS trisomic lines to determine necessity of Synj1 for aberrant phenotypes. Further, it is highly likely that novel cellular targets will emerge from corresponding lipidomic and RNAseq studies of functional iPSC derived neurons. We will also develop critical human neuronal cell models amenable to future therapeutic development.